Minor modifications in obtainable Arabidopsis floral dip method enhances transformation efficiency and production of homozygous transgenic lines harboring a single copy of transgene

Abstract

Many researchers have developed various methods for in-planta or floral dip transformation of Arabi-dopsis thaliana, one of the simple protocol and widely used to produce transgenic Arabidopsis. As the effi-ciency and ease of getting a transformant is very much time consuming effort and less number of the transformants people get, we have developed a little modified transformation protocol to avoid the dis-parities. Four types of inoculums (inoculum-1, in-oculum-2, inoculum-3 and inoculum-4) were used to check the transformation efficiency out of which In-oculum-3 showed the highest rate of transformation among the four types. 0.07% Twin-20 also acts in same manner as silwet L-77 to increase the rate of transformation efficiency and glucose instead of su-crose can be used in inoculum to transform Arabi-dopsis. After vacuum infiltration keeping the Agro-bacterium infected plants for 7 -8 hrs horizontally in low light at 28 0 C temperature condition, considered best to get an increased number of transformed seeds. Modified protocol produced ~ 12% -14% increase in transformants. Selection pots (kanamycin supple-mented soil filled pots) in place of selection plates (Kanamycin supplemented Murashige and Skoog agar plates) proved beneficial as no MS medium and no aseptic condition is required for selection of transformed plants. This increase in transformation efficiency consequently increased the percentage of homozygous and single copied stable transgenic lines.